Resolution of amino acids



United States Patent 2,790,001 RESOLUTION OF AMINO ACIDS Joseph L.Purvis, Northbrook, Ill., assignor to Interna-- No Drawing. ApplicationNovember 30, 1954, Serial No. 472,211

12 Claims. (Cl. 260-534) This invention relates to a process forresolving racemic modifications of optically active compounds, and moreparticularly, to the resolution of racemic modifications of alpha-aminocarboxylic acids.

Glutamic acid and aspartic acid are well known optically activecompounds. However, only L-glutamic acid and L-aspartic acid are usefulat the present time, there being no known utility for D-glutamic acidand D-aspartic acid or their racemic modification; namely, DL-glutamicacid or DL-aspartic acid. Chemical processes for the synthesis ofglutamic acid and aspartic acid, however, result in the formation ofracemic modifications of these compounds, that is, optically neutralDL-glutamic acid and DL-aspartic acid, respectively. These racemicmodifications, although having no value in commerce as such,

production of synthetic L-glutamic acid and L-aspartic acid has been thelack of a commercially feasible procedure for resolving racemicmodifications of glutamic acid and-aspartic acid into their respectiveenantiomorphs.

One object of the instant invention is to provide a process for theresolution of DL-alpha-amino carboxylic acids.

Another object of the instant invention is to provide a process for theresolution of DL-glutamic acid.

Another object of this invention is to provide a process for theresolution of DL-aspartic acid.

Another object of this invention is to provide a process for theresolution of DL-leucine.

A further object of this invention is to provide a commercially feasibleprocess for the resolution of DL-alphaamino carboxylic. acids which donot require the use of expensive reagents or equipment.

In accordance with this invention, a racemic modification of analpha-amino carboxylic acid, such as, for example, aspartic acid,glutamic acid, or leucine, is resolved by preparing a solutioncontaining the racemic modification and an optically active form ofanother alpha-amino carboxylic acid, and separating solids from thesolution by crystallization. The crystals thus separated comprise aquantity of an optically active form of the racemic modificationoriginally in the solution and a quantity of the racemic modification inthe unresolved state with perhaps some of the optically activecform ofthe resolving compound; The optically active form of the racemicmodification being subjected to the resolution process may be separatedfrom the other compounds by standard procedures.

It is preferred to apply the process of this invention to the resolutionof DL-alpha-amino carboxylic acids containing between about four andabout nine carbon atoms and containing less than three carboxy groups,using as the resolving compound an optically active form of analpha-amino carboxylic acid containing between about four and about ninecarbon atoms and containing less in the molecule.

than three carboxy groups. Alpha-amino carboxylic acids which areparticularly useful in this invention are those having a solubility inwater at room temperature of less than about 8 grams per 100 grams ofwater. Specific alpha-amino carboxylic acids, in addition to leucine,aspartic acid and glutamic acid, which may be utilized include tyrosine,isoleucine, norleucine, valine, and the like. All of these compounds maybe utilized as resolving compounds or may be resolved when in theirracemic modifications according to the process of this invention. All ofthese compounds are characterized as non-basic alpha-amino carboxylicacids for the reason that all contain not more .than one amino group percarboxy group Alpha-amino carboxylic acids containing more than oneamino group per carboxygroup in the molecule, such as lysine, are notoperable in this invention.

"A preferred practice of the instant invention comprises preparing anaqueous solution of a racemic modification of either aspartic acid orglutamic acid, the solution also containing an optically active form ofthat one of these two compounds which is not being resolved. Forexample, if it is desired to resolve DL-aspartic acid in accordance withthis invention, a solution is prepared containing DL-aspartic acid andan optically active form of, for example, glutamic acid, that is, eitherL-glutamic acid or D-glutamic acid. 'Alternatively, if it is desired toresolve DL-glutamic acid in accordance with this invention, a solutionis prepared containing DL-glutamic acid and an optically 'active form ofaspartic acid, that is, D-' aspartic acid or L-aspartic acid.

A convenient method for preparing the solution'com prises preparing anaqueous slurry of the racemic modification and an optically active formof the resolving com-' pound, for example, an aqueous slurry ofDL-aspartic acid and L-glutamic acid, and adding alkali to the slurry insufiicient amount to dissolve the solids and form a true solution. Thesolution is then adjusted to about the isoelectric point of the racemicmodification by the addition of an inorganic acid, such as aqueoushydrochloric acid, and the volume of the solution is adjusted by addingor removing water so that the solution will be supersaturated withrespect to the racemic modification at the isoelectric point. tocrystallization contain between about 3% and about 5% of the racemicmodification and between about 1% and about 2% of the resolvingcompound. The solution is then allowed to stand with gentle agitationuntil crystallization is complete. Crystallization is usually carriedout at room temperature, but higher or lower temperatures may beutilized if desired. The crystals will cont-ain a substantial quantityof an optically active form of of the racemic modification correspondingin rotation to that of the resolving compound, and also a substantialquantity of the racemic modification.

-In accordance with a specific embodiment ofthis invention, DL-asparticacid and L-glutamic acid are slurried in water, and aqueous sodiumhydroxide is added to the slurry with agitation until the pH of theliquid phase is adjusted, preferably to about 7.0. Under theseconditionsv the DL-aspartic acid and L-glutamic acid dissolve to form atrue solution. The solutionis then adjusted to about.

. pH 3.1 by'the addition of concentrated aqueous hydrochloric acid, andthe volumeof the solution is adjusted It is desirable that the solutionbeing subjected armour by adding or removing (by evaporation) water sothat the solution at pH 3.1 will be supersaturated with respect toDL-aspartic acid. The solution is then allowed to stand with gentleagitation for about 72 hours at room temperature and filtered. Thecrystals obtained comprise about 45% DL-aspartic acid, about 40% ofD-aspartic acid, and about 15% QtL-gIutamic acid. The filtrate containsabout 63% of the DL-aspart-ic acid originally in solution, about 8%L-aspartic acid (based upon total solids in solution) and about 90% ofthe L-glutamic acid originally added.

In carrying out the process of this invention, it is desirable that thecrystallization be carried out at a temperature of about roomtemperature or below, but higher temperatures may be utilized forparticular purposes. The solution of the racemic modification and theresolving compound may be prepared in any convenient manner so long asthe solution upon adjustment to about the isoelectric point of theracemic modification is supersaturated with respect to the racemicmodification. It is not necessary that the resolving compound be atits-saturation concentration at the time of crystallization if less thanoptimum resolution is permissible. The best yields, however, areobtained if the resolving compound is at its saturation concentration atthe time that crystallization begins. Crystallization is carried outuntil the formation of crystals discontinues, that is, until equilibriumconditions obtain or are substantially achieved.

The term resolving compound as used herein refers to an optically activeform of either glutamic acid or aspartic acid, or similar non-basicalpha-amino carboxylic acid, for example, L-aspartic acid, D-asparticacid, L-glutamic acid, or D-glutamic acid, etc. When DL-aspartic acid issubjected to resolution in accordance with this invention, the resolvingcompound is preferably either L-glutamic acid or D-glutamic acid.Conversely, when DL-glutamic acid is resolved in accordance with thisinvention, the resolving compound is preferably L-aspartic acid or D-aspartic acid.

, T he following examples representspecifieembodiments ofthis invention.Allparts and percentages are by weight unless otherwise indicated.

Example I About 1'0'parts of DL-asparatic acid and about '5 parts ofL=glutamic acid were slurried with about 200 parts water. To the slurrywas added a 50% aqueous of sodium hydroxide until the 'pH of the aqueousphase amounted to about 7.0. Addition of the sodium hydroxide solution"resulted in the dissolution of the DL-aspartic acid and L-glutamic acidto form a true solution. The solution was adjusted to pH 3.1 by theaddition of 37% aqueous hydrochloric acid and then sufficient water wasadded to the solution to adjust the weight of the solution to 250parts.The solution was allowed to stand with gentle agitation for about 72hours at room temperature and then filtered to remove the crystals whichhad formed. The crystals amounted to 5.8 parts and had the followingcomposition by weight:

Percent D-aspartic acid 37.0 DL-aspartic acid. 54.5 L-glutamic acid 8.5

The .filtrate amounted to 240 parts and contained :the following solids:

Parts L-aspartic acid--. 2.1 DL-aspartic acid 2.6 Lvglutamic acid 4.4

glutamic acid by re-crystallizing the D-aspartic acid at itsi'soelectric point (pH 3.1).

The L-aspartic acid was isolated from the filtrate by first separatingL-glutamic acid by crystallization of L- glutamic acid hydrochloride,adjusting the pH to 3.1 and erystallizing L-aspartic acid.

Example 11 Twelve parts DL-aspartic acid and 8 parts L-glutamic acidwere slurried in Water and dissolved by adjusting the pH of the aqueousphase to about 7.0 by the addition of a 50% aqueous solution of sodiumhydroxide. The solution was then adjusted to pH 3.2 by the addition of30% aqueous hydrochloric acid, and the weight of the solution was thenadjusted to 400 parts by adding water. The solution was allowed to standwith gentle agitation for about 72 hours and then filtered to remove thecrystals which had formed. The crystals amounted to 3.5 parts and hadthe following compositions:

Percent D-aspartic acid 40 DL-aspartic acid 45 L-glutamic acid 15 Thefiltrate amounted to 393 parts and contained the following solids:

Parts L-aspartic acid 1.4 DL-aspartic acid 7.6 L-glutamic acid 7.2

The optically active forms of aspartic acid and glutamic acid wererecovered by the procedures outlined in Example I.

Example III About 10 parts of DL-glutamic acid and about 5 parts ofL-aspartic acid were slurried in about 200 parts water. To the slurrywas added a. 5 0% aqueous solution of sodium hydroxide until the pH ofthe aqueous phase amounted to about 7.0. Addition of the sodiumhydroxide solution resulted'in dissolution of the DL-glutamic acid andL- aspartic acid to form a true solution. The solution was adjusted topH 3.2 by the addition of 37% aqueous hydrochloric acid and thensuificient water was added to the solution to adjust the weight of thesolution ;to 250 parts. The solution was allowed to stand with gentleagitation for about 72 hours at room temperature and then filtered toremove the crystals which had formed. The crystals amounted to about 5parts and had the following composition by weight:

Percent D-glutarnic acid 10 DL-glutamic acid 78 L-aspartic acid 12Example TV About 16.8 parts of DL-glutamic acid monohydrate and about 9parts of L-leucine were dissolved in dilute hydrochloric acid at pH 1.0.The solution was :then adjusted to pH 3.2 by the addition of a 50%aqueous sodium hydroxide solution. The final solution amounted to about300 parts. The solution was agitated gently over a period of about24'hours at about 25 C. to per mit crystallization to proceed, and thecrystals were re moved by filtration. The crystals amounted to 9.4 partsand comprised 1.8 parts D-glutamic acid and 7.6 parts DL-glutamic acid.The Drglutamic acid was separated from the .DL-glutamic acid byrecrystallization from 300 parts of water at about 25 C.

Having thus fully described and illustrated the character of theinstantinvention, What is desired to be secured and claimed by LettersPatentis:

1. A process forresolvinga racemic-modification of :a non-basicalpha-amino 'carboxylic acid comprising prc paring a solution containingsaid racemic modification and an optically active .form of anothernon-basic alphaamino carboxylic acid, and separating solids comprisingan optically active form of the racemic modification from the solutionby crystallization.

2. A process for resolving a racemic modification se lected from thegroup consisting of DL-aspartic acid and DL-glutamic acid, comprisingpreparing a solution containing one of said racemic modifications and anoptically active form of the other, and separating solids comprising anoptically active form of the racemic modification from the solution bycrystallization.

3. A process for resolving a racemic modification selected from thegroup consisting of DL-aspartic acid and DL-glutamic acid, comprisingpreparing an aqueous solution supersaturated with one of said racemicmodifications, and containing an optically active form of the other, andseparating solids comprising an optically active form of the racemicmodification from the solution by crystallization.

4. A process for resolving DL-aspartic acid which comprises preparing anaqueous solution supersaturated with respect to DL-aspartic acid andalso containing L-glutamic acid, and separatin solids comprising anoptically active form of aspartic acid from the solution bycrystallization at pH 3.1.

5. A process for resolving DL-aspartic acid which comprises preparing anaqueous solution supersaturated with respect to DL-aspartic acid andsaturated with L-glutamic acid, and separating solids comprising anoptically active form of aspartic acid from the solution bycrystallization at pH 3.1.

6. A process for resolving DL-aspartic acid which comprises preparing anaqueous solution supersaturated with respect to DL-aspartic acid andalso containing D-glutamic acid, and separating solids comprising anoptically active form of aspartic acid from the solution bycrystallization.

7. A process for resolving DL-aspartic acid which comprises preparing anaqueous solution supersaturated with respect to DL-aspartic acid andsaturated with D-glutamic acid, and separating solids comprising anoptically active form of aspartic acid from the solution bycrystallization.

8. A process for resolving DL-glutamic acid which comprises preparing anaqueous solution supersaturated with respect to DL-glutamic acid andalso containing L- aspartic acid, and separating solids comprising anoptically active form of glutamic acid from the solution bycrystallization at pH 3.2.

9. A process for resolving DL-glutamic acid which comprises preparing anaqueous solution supersaturated with respect to DL-glutamic acid andsaturated with L- aspartic acid, and separating solids comprising anoptically active form of glutamic acid from the solution bycrystallization at pH 3.2.

10. A process for resolving DL-glutamic acid which comprises preparingan aqueous solution supersaturated with respect to DL-glutarnic acid andalso containing D-aspartic acid, and separating solids comprising anoptically active form of glutamic acid from the solution bycrystallization.

11. A process for resolving DL-glutamic acid which comprises preparingan aqueous solution supersaturated with respect to DL-glutamic acid andsaturated with D- aspartic acid, and separating solids comprising anoptically active form of glutamic acid from the solution bycrystallization.

12. A process for resolving DL-glutarnic acid which comprises preparingan aqueous solution supersaturated with respect to DL-glutamic acid andcontaining an optically active form of leucine, and separating solidscomprising an optically active form of glutamic acid from the solutionby crystallization.

References Cited in the file of this patent UNITED STATES PATENTS Emmicket al Jan. 2, 1951 Wiedman July 13, 1954 OTHER REFERENCES

1. A PROCESS FOR RESOLVING A RACEMIC MODIFICATION OF A NON-BASICALPHA-AMINO CARBOXYLIC ACID COMPRISING PREPARING A SOLUTION CONTAININGSAID RACEMIC MODIFICATION AND AN OPTICALLY ACTIVE FORM OF ANOTHERNON-BASIC ALPHAAMINO CARBOXYLIC ACID, AND SEPARATING SOLIDS COMPRISINGAN OPTICALLY ACTIVE FORM OF THE RACEMIC MODIFICATION FROM THE SOLUTON BYCRYSTALLIZATION.